The growth of biological research has resulted in a number of assay techniques such as reporter gene assays, new pharmaceutical compound screening, high-throughput polymerase chain reaction (“PCR”), real-time PCR, and similar assays, and has created a need for handling large numbers of test samples at one time in order to control costs and efficiently handle these large numbers of samples. A number of analytical methods are now available for high-throughput screening of these samples. One important method for monitoring samples in high-throughput setting is the use of luminescence from fluorescent dyes and the like. Such substances can be used as a tag, reporter, or marker for various test assays where luminescence can be used as a direct or indirect measure of a selected property in a sample well.
In a high-throughput or real-time assay, large numbers of samples are typically processed and analyzed together (e.g., by luminescence emission) in a multi-well sample plate called an assay microplate. These microplates provide an array of wells, usually 48 or 96 wells in typical examples, but 384 well and 1536 well microplates are becoming more common as well. The most common configuration at this time is the 96-well format (in an 8 by 12 configuration). Illustratively, all plates, regardless of the number of wells, can have roughly the same dimensions. It is also possible to have plates of other configurations and dimensions.
In many assays, assay microplate wells are filled with test samples and then placed in an instrument that may include a detector system, such as a luminescence microplate reader for measuring the relative luminescence emissions of each test well. Since different luminescence materials used for microplates assays produce different degrees of light intensity, light intensity can act as a direct or indirect measure of test results—i.e., the greater the light intensity, the greater the result. For example, some PCR thermocyclers include optical systems for real-time monitoring of luminescence emission of samples undergoing thermocycling for nucleic acid amplification or for post-amplification melting. In such systems, fluorescent emission of one or more reagents included in the sample(s) can be used to monitor things such as template concentration, product concentration, DNA melting point temperature, and the like.
Although luminescence microplates and luminescence microplate readers and similar instruments are of great utility in automated screening, there are a number of issues that affect their use. In particular, unless an optical device (e.g., a digital or analog camera or another detector) is moved sufficiently far from the microplate, the optical device cannot typically see to the bottom of all of the wells simultaneously. This is illustrated in FIG. 1, which shows a view of a 96-well plate 100 from a relatively close distance. As can be seen in FIG. 1, a camera or another type of detector can see all the way to the bottom of the wells only in the center of the plate in the region labeled 110. However, because of obstruction from the walls of the wells, the camera cannot see into the bottom of the wells in the region outside region 110. This is illustrated, for example, at wells 120a and 120b. A number of systems have been developed to allow microplate readers and similar instruments to see into all of the wells of a microplate.
In one example, the detector (e.g., an optical device, such as a digital camera) can be moved far enough from the microplate to allow the detector to see to the bottom of all of the wells. However, moving the optical device sufficiently far from the microplate increases the form factor of the plate reader and typically makes the plate reader large and cumbersome. Likewise, arbitrarily increasing the distance between the optical device and the microplate leads to diffusion of light and loss of signal from the microplate and can also lead to cross talk between signals from adjacent samples wells.
In another example, a luminescence microplate reader may have a series of optical devices wherein each device is positioned to correspond to a well in the microplate that holds a test sample. The optical device is often a photodiode.
In yet another example, the luminescence microplate reader may be fitted with a single optical device wherein the plate, the reading device, or both are moved sequentially to the appropriate reading position in order to detect the luminescence in each well. Alignment can be adversely affected when one or more of the aligned components involved is shifted in position or becomes damaged. If, for any reason, the alignment is incorrect, the wells will not be centered properly in alignment with the optical reading device, resulting in an incorrect luminescence measurement
In still yet another example, the luminescence microplate reader may be fitted with a single optical device and one or more optical lenses that focus the signal from the wells onto the optical device. However, such optical lens systems can be expensive and cumbersome. Moreover, the optics used to measure luminescence must avoid cross-talk from one sample well when measuring the luminescence from an adjacent well.